UC San Diego

Hexokinase-II (HK-II), the first glycolytic kinase, is found not only in the cytosol, but also at mitochondria. We previously showed that interventions increasing mitochondria associated HK-II are cardioprotective. In this study, we tested the hypothesis that mitochondrial HK -II regulates mitochondria specific autophagy (mitophagy), removing damaged mitochondria by lysosomal degradation to preserve cellular integrity. We showed that mitochondrial HK-II is significantly decreased in the whole heart as well as neonatal rat ventricular myocytes (NRVMs) when subjected to ischemia. To explore the consequences of mitochondrial HK-II dissociation, a dissociation peptide (15G) was expressed in NRVMs. Expression of 15G decreased mitochondrial HK-II by 40%. Peptide treatment also increased mitochondrial Parkin, an important mitophagic ubiquitin ligase, as well as ubiquitination of mitochondrial proteins and decreased expression of the mitochondrial protein VDAC. Expression of 15G did not decrease cytosolic or nuclear proteins, nor did it induce mitochondrial membrane depolarization. The VDAC decrease is recovered by inhibition of lysosome activity, suggesting that HK-II dissociation regulates lysosome dependent mitophagy. PINK1, a mitochondrial kinase implicated in Parkin recruitment, was not observed, suggesting that mitochondrial HK-II dissociation recruits Parkin through a mechanism distinct from the established role of PINK1 in Parkin recruitment to depolarized mitochondria. Mitofusin2 (mfn2) has been shown to be a mitochondrial Parkin receptor. Our study revealed that HK- II binds to mfn2 and that this binding is diminished by 15G expression or ischemia. Together these results suggest that HK-II dissociation plays a regulatory role in Parkin translocation and induction of mitophagy in response to ischemia in cardiomyocytes